1. An Essential Role for IFN-γ in Regulation of Alloreactive CD8 T Cells Following Allogeneic Hematopoietic Cell Transplantation 
Wannee Asavaroengchai, Hui Wang, Shumei Wang, Lan Wang, Roderick Bronson, Megan Sykes, Yong-Guang Yang 
Biology of Blood and Marrow Transplantation 
Volume 13, Issue 1, Pages (January 2007)
DOI: /j.bbmt
Copyright © 2007 American Society for Blood and Marrow Transplantation Terms and Conditions

2. Figure 1
IFN-γ inhibits GVHD induced by donor CD8 T cells. Recipient B6D2F1 mice were lethally irradiated and reconstituted with 7 × 106 TCD BMCs and 3 × 107 CD4-depleted spleen cells from WT (WT GVHD; n = 15) or GKO (GKO GVHD; n = 19) B6 donors. Non-GVHD controls were lethally irradiated B6D2F1 mice that received TCD BMC alone or with TCD splenocytes from WT (WT-Ctrl; n = 8) or GKO (GKO-Ctrl; n = 8) donors. Data from 3 independent experiments are combined. The percentages of CD8+ cells in WT and GKO allo-HCT inocula (ie, CD4-depleted splenocytes) used in these experiments were 17.9% and 17.7%, 16.1% and 15.7%, and 21% and 23%, respectively. Shown are (A) percent survival and (B) mean body weight changes (mean ± SDs) of the allo-HCT recipients. Mean weight changes = [(current weight/weight at day 0) − 1]. *P < .05.
Biology of Blood and Marrow Transplantation  , 46-55DOI: ( /j.bbmt )
Copyright © 2007 American Society for Blood and Marrow Transplantation Terms and Conditions

3. Figure 2
GKO allo-HCT induces more severe GVHD pathology. Shown are pathologic GVHD scores (mean ± SD, n = 3-6/group) and representative H&E photomicrographs of the lungs (A), liver (B), and colon (C) from the recipients of WT allo-HCT (solid columns) or GKO allo-HCT (open columns). *P < .05. D, Immunofluorescent staining of liver sections prepared from allo-HCT recipients at day 7 with anti-CD8. Three mice from each group were analyzed and representative photomicrographs are shown.
Biology of Blood and Marrow Transplantation  , 46-55DOI: ( /j.bbmt )
Copyright © 2007 American Society for Blood and Marrow Transplantation Terms and Conditions

4. Figure 3
Augmented activation and expansion of donor CD8 T cells in the recipients of GKO allo-HCT. A, B, Expression of activation markers on donor CD8 T cells in the spleens of lethally irradiated B6D2F1 mice receiving allo-HCT from WT or GKO B6 donors (n = 6/group). A, Percentages of donor CD25+, CD62L−, and CD44high CD8 T cells at the indicated time points. WT and GKO donor CD8 T cells are presented as solid and open bars, respectively. B, Representative flow cytometric profiles display the expression of CD25, CD62L, and CD44 on gated donor CD8 T cells at day 4. Thick lines represent the isotype staining controls. Analysis gates were set based on staining profiles of appropriate isotype control mAs and naive splenocytes were stained with the same mAbs at the same time. C, Spleen cells were prepared from B6D2F1 recipients of allo-HCT from WT (solid bars) or GKO (open bars) 2C donors at the indicated time points, and the percentages of CD25+ and CD62L− 2C T cells were analyzed by flow cytometric analysis. Data are shown as mean ± SD (n = 3/group). *P < .05; **P < .01.
Biology of Blood and Marrow Transplantation  , 46-55DOI: ( /j.bbmt )
Copyright © 2007 American Society for Blood and Marrow Transplantation Terms and Conditions

5. Figure 4
Effect of IFN-γ on apoptosis of donor CD8 T cells. Spleen cells were prepared from the recipients of WT (solid bars) or GKO (open bars) allo-HCT at the indicated time points, and the percentages of apoptotic donor CD8 (or 2C) T cells were determined by flow cytometric analysis. A, Percentages of apoptotic donor CD8 T cells in the spleens of B6 → B6D2F1 HCT recipients. B, Percentages of apoptotic donor 2C T cells in the spleens of 2C →; B6D2F1 HCT recipients. Data are shown as mean ± SD (n = 3-6/group). *P < .05; **P < .01.
Biology of Blood and Marrow Transplantation  , 46-55DOI: ( /j.bbmt )
Copyright © 2007 American Society for Blood and Marrow Transplantation Terms and Conditions

6. Figure 5
Effect of IFN-γ on division of donor CD8 T cells. A, B, B6D2F1 mice received BMCs and CSFE-labeled CD4-depleted spleen cells from WT (solid bars) or GKO (open bars) B6 donors (n = 3/group). Recipient spleen and lymph node cells were prepared at the indicated time points and analyzed for division of donor CD8 T cells. A, Numbers of donor CD8 T cells with each division in the spleen at days 2 and 4 and in the lymph nodes at day 4 after HCT. Axillary, brachial, and inguinal lymph nodes were harvested at the indicated time points, and pooled lymph node cells were analyzed. B, Representative histograms show CFSE levels in gated donor CD8 T cells at day 4. Numbers indicate percentages of cells with each number of cell divisions. C, B6D2F1 mice received BMCs and CFSE-labeled spleen cells from WT (solid bars) or GKO (open bars) 2C donors (n = 3/group). Data shown are percentages of donor 2C TCR+ cells with different cell divisions (mean ± SD) in the recipient spleen at day 4 after HCT. *P < .05; **P < .01; ***P < .001.
Biology of Blood and Marrow Transplantation  , 46-55DOI: ( /j.bbmt )
Copyright © 2007 American Society for Blood and Marrow Transplantation Terms and Conditions

7. Figure 6
Comparison of Treg, NKT, and memory T cells in normal WT and GKO B6 mice. Spleen cells were prepared from naive WT (solid bars) and GKO (open bars) B6 mice (8 wk old; n = 3/group) and analyzed for Treg, NKT, and memory T cells by flow cytometry. A, Percentages of total (CD3+Foxp3+), CD4 (CD4+Foxp3+), CD8 (CD8+Foxp3+), and CD4− (CD4−Foxp3+) Treg cells. B, Percentages of total (CD3+NK1.1+), CD4 (CD4+CD3+NK1.1+), CD8 (CD8+CD3+NK1.1+), and double-negative (CD4−CD8−CD3+NK1.1+) NKT cells. C, Percentages of CD4 and CD8 central (CD44+CD62L+) and effector (CD44+CD62L−) memory T cells. Results are presented as mean ± SD. *P < .05; **P < .01.
Biology of Blood and Marrow Transplantation  , 46-55DOI: ( /j.bbmt )
Copyright © 2007 American Society for Blood and Marrow Transplantation Terms and Conditions